This invention relates to the spectral analysis of small amounts of substances deposited on a substrate, which is maintained at a uniform low temperature (in an evacuated chamber). In particular, it concerns the use of infrared spectroscopy in the analysis of chromatographic fractions.
Chromatographic analysis is significantly improved by depositing (and freezing) the separated fractions on a substrate, and using infrared spectroscopy (FTIR) for sample analysis. In Reedy U.S. Pat. No. 4,158,772, issued June 19, 1979 various prior types of chromatographic analysis are discussed (Col. 1, lines 55-67), which "exhibit limited observation time and consequently suffer in both precision and resolution". The Reedy patent discloses a "matrix-isolation" technique, in which separated fractions are "entrapped within a frozen matrix of an inert substance such as argon or krypton gas".
The apparatus disclosed in the Reedy patent requires cooling the substrate to a very low temperature, e.g., the temperature provided by liquid helium. Maintenance of the required extremely low temperature requires very expensive apparatus.
A less costly apparatus for analysis of chromatographic fractions has been proposed in an article by Pentony, Shafer & Griffiths in the June, 1986 issue of the Journal of Chromatographic Science. They describe a system for analysis of chromatographic fractions, which involves depositing the fractions in a narrow track on a cooled transparent substrate, and then analyzing the deposited material by means of an infrared transmission microscope attached to an FTIR spectrometer. This system represents an advance over other means for the IR analysis of chromatographic fractions (whether gas, liquid, or supercritical fluid). However, it does have some drawbacks. In particular, the need to analyze the deposit in transmission (i.e., radiation entering on one side and exiting on the other) necessitates cooling the substrate from the edges. This tends to limit temperature uniformity, particularly at low temperatures such as liquid nitrogen temperature (77 degrees Kelvin). In addition, the need to focus the radiation to a small spot requires the use of optical elements quite close to the substrate, thus restricting the space available for cooling, for vacuum apparatus, and for the needed computer-controlled positioning devices.
It has been recognized that these space constraints could be relieved if the fractions could be deposited on a reflecting substrate and analyzed by means of reflectance microscopy. This would leave all the space on one side of the substrate free for cooling, and for the other required apparatus. However, experiments thus far have established that the spectra obtained in this way contained undesired artifacts, probably due to large spectral variations in the amount of radiation reflected from the surface of the deposited material. These variations are related to differences in its index of refraction (generally referred to as dispersion).